Chromatography system and method

ABSTRACT

A chromatography system and method for analyzing a sample of interest. The chromatography system can include a housing, a cartridge holder coupled to the housing and formed to hold a chromatography cartridge containing a stationary phase, a collection vessel stand holder coupled to the housing and formed to hold a collection vessel stand, and a pump coupled to the housing. A fluid path can be coupled to the housing and can connect the pump to the cartridge holder to direct a mobile phase from the pump to the cartridge holder and can further connect the cartridge holder to the collection vessel stand holder to direct the mobile phase from the cartridge holder to the collection vessel stand holder. The chromatography system can further include a detector positioned along the fluid path between the cartridge holder and the collection vessel stand holder, and a controller integrally coupled to the housing.

RELATED APPLICATIONS

This application is a divisional of U.S. patent application Ser. No.11/084,741, filed on Feb. 25, 2005, which claims priority to U.S.Provisional Patent Application No. 60/547,613, filed on Feb. 25, 2004,the entire contents of which are incorporated herein by reference.

BACKGROUND

The present invention relates generally to a chromatography system andmethod, and particularly, to a liquid chromatography system and method,and more particularly, to a flash chromatography system. Chromatographyis used to analyze the constituents, or fractions, of a sample ofinterest, and, in some cases, to collect each of the fractions of thesample of interest separately for further analysis or use.Chromatography generally relates to any of a variety of techniques usedto separate complex mixtures based on the differential affinities of thefractions of the sample for a mobile phase with which the sample flows,and a stationary phase through which the sample passes.

Generally, liquid chromatography includes a stationary phase thatincludes a finely powdered solid adsorbent packed into a chromatographycartridge or column, and the mobile phase includes one or more elutingsolvents that are moved through the cartridge by a pump. The sample tobe analyzed by liquid chromatography is injected into the cartridge andmonitored by a detector. The detector provides identification and/ordifferentiation of the fractions as the fractions elute from thecartridge. In general, flash chromatography includes a cartridge (insome cases, a disposable cartridge) filled with the stationary phase(e.g., silica gel), and the sample to be separated is placed on top ofthe stationary phase. The cartridge is filled with an isocratic orgradient solvent which, with the help of pressure, enables the sample torun through the cartridge and become separated. Generally, liquidchromatography, and particularly, flash chromatography can be used for avariety of applications, including, but not limited to, drug discovery,sample clean-up, and natural product purification, among others.

SUMMARY

Some embodiments of the present invention provide a chromatographysystem for analyzing a sample of interest, and the sample includingfractions. The chromatography system can include a housing; a cartridgeholder coupled to the housing and formed to hold a chromatographycartridge containing a stationary phase; a collection vessel standholder coupled to the housing and formed to hold a collection vesselstand; a fraction collector coupled to the housing, the fractioncollector selectively dispensing fractions to the collection vesselstand holder; a pump coupled to the housing; a fluid path coupled to thehousing and connecting the pump to the cartridge holder to direct amobile phase from the pump to the cartridge holder and connecting thecartridge holder to the collection vessel stand holder to direct themobile phase from the cartridge holder to the collection vessel standholder; a detector positioned along the fluid path between the cartridgeholder and the collection vessel stand holder; and a controllerintegrally coupled to the housing, the controller controlling the pumpand receiving data from the detector.

Other features and aspects of the invention will become apparent byconsideration of the detailed description, accompanying drawings, andclaims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a front perspective view of a chromatography system accordingto one embodiment of the present invention.

FIG. 2 is a right rear perspective view of the chromatography system ofFIG. 1.

FIG. 3 is a left rear perspective view of the chromatography system ofFIGS. 1-2.

FIG. 4 is a front elevational view of the chromatography system of FIGS.1-3.

FIG. 5 is a side elevational view of the chromatography system of FIGS.1-4.

FIG. 6 is a top view of the chromatography system of FIGS. 1-5.

FIG. 7 is a perspective view of a pump assembly according to oneembodiment of the present invention.

FIGS. 8-20 illustrate various screen shots of a graphical user interfaceaccording to one embodiment of the present invention.

FIG. 21 is a schematic view of a touchpad according to one embodiment ofthe present invention.

DETAILED DESCRIPTION

Before any embodiments of the invention are explained in detail, it isto be understood that the invention is not limited in its application tothe details of construction and the arrangement of components set forthin the following description or illustrated in the following drawings.The invention is capable of other embodiments and of being practiced orof being carried out in various ways. Also, it is to be understood thatthe phraseology and terminology used herein is for the purpose ofdescription and should not be regarded as limiting. The use of“including,” “comprising,” or “having” and variations thereof herein ismeant to encompass the items listed thereafter and equivalents thereofas well as additional items. Unless specified or limited otherwise, theterms “mounted,” “connected,” “supported,” and “coupled” and variationsthereof are used broadly and encompass both direct and indirectmountings, connections, supports, and couplings. Further, “connected”and “coupled” are not restricted to physical or mechanical connectionsor couplings. Furthermore, terms such as “front,” “rear,” “top,”“bottom,” and the like are only used to describe elements as they relateto one another, but are in no way meant to recite specific orientationsof the apparatus, to indicate or imply necessary or requiredorientations of the apparatus, or to specify how the invention describedherein will be used, mounted, displayed, or positioned in use.

FIGS. 1-6 illustrate a chromatography system 100 according to oneembodiment of the present invention. The chromatography system 100 canbe used to detect and separate a sample 101 into its constituents, orfractions. The chromatography system 100 includes a housing 102; acontroller 104 with chromatography programming software having agraphical user interface 106; a user interface 108 to allow a user tocontrol various aspects of the system 100; a pump assembly 110 includinga pump 114 for moving a mobile phase comprising one or more solvents 115through a fluid path 112 in the chromatography system 100, and, in someembodiments, a mixing valve 113 for mixing the one or more solvents 115;a sample injector 116; a cartridge holder 118 for holding achromatography cartridge 120 into which the sample 101 will be injectedand through which the sample 101 will pass to be separated; a detector122 for detecting the fractions of the sample 101; and a fractioncollector 124 for ejecting fractions of the sample 101, or the entiresample 101, from the chromatography system 100 into one or morecollection vessels 126. In some embodiments, the collection vessels 126are positioned in one or more collection vessel stands 128. Thecollection vessels 126 can include a variety of containers, including,without limitation, at least one of test tubes 126, as shown in FIGS.1-6, arranged in one or more test tube racksl28, micro plates, microvials, scintillation vials, etc.

As used herein and in the appended claims, the term “fluid path” 112refers collectively to those areas in the chromatography system 100through which fluid passes. The “fluid path” 112 can include an entirepath followed by fluid through the chromatography system 100 or canrefer to a portion of that path.

As used herein and in the appended claims, the terms “upstream” and“downstream” refer to the direction of fluid movement in thechromatography system 100. That is, the term “upstream” is used todescribe any location, element or process that occurs prior to the pointor area being referred to relative to the direction of fluid movement inthe chromatography system 100, whereas the term “downstream” is used todescribe any location, element or process that occurs subsequent to thepoint or area of reference with respect to fluid movement in thechromatography system 100.

In general, the sample 101 can be loaded directly into an inlet 119 ofthe cartridge 120, or the sample 101 can be loaded into the sampleinjector 116 to be dissolved (e.g., in a stronger solvent to reducesample viscosity), and then injected into the inlet 119 of the cartridge120. After the sample 101 has been added to the cartridge 120, thesample 101 interacts with the mobile phase of the system 100 and astationary phase 121 contained within the cartridge 120. The mobilephase and the sample 101 are moved by the pump 114 through the cartridge120 at a particular flow rate, or programmed sequence of flow rates.Based on the varying affinities of the fractions of the sample 101 forthe solvents 115 and the stationary phase 121, the fractions of thesample 101 will pass through the stationary phase 121 at different flowrates, and thus, elute from an outlet 123 of the cartridge 120 atdifferent times. Each fraction will then flow from the cartridge 120 tothe detector 122 positioned downstream from the cartridge 120 in thefluid path 112, and to the fraction collector 124 to be expelled intoone or more test tubes 126.

The housing 102 of the illustrated embodiment is formed of severalportions that each house and/or support the various components of thesystem 100. In some embodiments, the portions of the housing 102 areintegrally formed, and the housing 102 is a unitary body. In someembodiments, the portions of the housing 102 are each formed ofdifferent parts. For example, in the embodiment illustrated in FIGS.1-6, the housing 102 is formed of separate parts that are coupledtogether in a stacked configuration to position the various componentsof the system 100 in a substantially vertical arrangement. The verticalarrangement of the system 100 creates a relatively compact system 100that requires a relatively minimal amount of surface area or workspaceon a desk, table or lab bench. In addition, the housing 102 allows thecomponents of the system 100 to be positioned in relatively closeproximity to one another and to be coupled together to form anintegrated chromatography system 100. The integrated chromatographysystem 100 includes substantially all of the necessary components anddevices to perform chromatography, and thus allows for facileinstallation, set-up and use. In addition, the integrated chromatographysystem 100 is sized and configured such that substantially the entirechromatography system 100 can be positioned on a lab bench, inside achemical hood, and the like. The housing 102, or portions thereof, canalso be used to protect or conceal (e.g., for aesthetic purposes)various portions of the chromatography system 100. For example, in someembodiments, as shown in FIG. 7, the pump assembly 110 is positioned ina base portion 127 of the housing 102, and is concealed from view by afront plate 129, as shown in FIGS. 1-6.

The housing 102 illustrated in FIGS. 1-6 is shown by way of exampleonly, and it should be understood that other housing configurations anddesigns can be used to form a relatively compact and integratedchromatography system 100. In addition, it should be understood that thehousing 102 can instead be formed of a unitary body. Furthermore, theorganization and arrangement of the components of the chromatographysystem 100 is shown by way of example only, and a variety of otherarrangements of the components of the chromatography system 100 can beused to form the integrated chromatography system 100, without departingfrom the spirit and scope of the present invention.

The controller 104 is internal to the chromatography system 100, and isintegral with the chromatography system 100 to eliminate the need for anindependent, external personal computer. The controller 104 controlsvarious aspects of the chromatography process. In some embodiments, thecontroller 104 of the chromatography system 100 is microprocessor-basedand is adapted to allow the user to interact with and manipulate thecomponents of the chromatography system 100 to perform a chromatographyanalysis of the sample 101 of interest. For example, in the embodimentillustrated in FIGS. 1-6, the controller 104 includes computer controlhardware with an embedded Intel® Strong ARM 206 MHz RISC processor and aMICROSOFT® Windows CETm-based operating system, such as INTELLIPEAK™chromatography programming software, available from Analogix, Inc.,Burlington, Wis. In other embodiments, the controller 104 can be anyprogrammable or non-programmable electronic system, and need notnecessarily be microprocessor-based. The controller 104 can include anycombination of hardware and/or software components. By way of exampleonly, the controller 104 can include any number of discrete logicelements coupled together to perform the same function as describedabove. Still other types of electronic controllers capable of performingthis function are possible, would be readily recognized by those skilledin the art, and fall within the spirit and scope of the presentinvention.

In some embodiments, as shown in the embodiment illustrated in FIGS.1-6, the controller 104 includes standard personal computer hardware andsoftware. For example, in the embodiment illustrated in FIGS. 1-6, thecontroller 104 includes MICROSOFT® Windows® CE desktop software. Thecontroller 104 can be connected to one or more external controllers(e.g., personal computers) directly or via a network (e.g., a local areanetwork (LAN) or the internet) for data transfer. For example, thecontroller 104 can be connected to the internet wirelessly or via anEthernet connection for data transfer.

The controller 104 can also include a variety of standard input jacks,including a universal serial bus (USB), which can allow a variety ofexternal devices to be connected to the controller 104. For example,memory storage devices can be connected to the controller 104 via a USBconnection for file transfer. The controller 104 can include a varietyof standard programs (e.g., MICROSOFT® EXCEL™ spreadsheet program) fordata storage and analysis, word processing, and the like. The controller104 can further include a variety of drive devices for file storage andtransfer, including, without limitation, at least one of a floppy diskdrive, a CD-ROM drive, a DVD-ROM drive, a CD-R drive, a DVD-R drive, aCD-RW drive, a zip drive, and the like. In some embodiments, thecontroller 104 can include a compact flash memory drive device. In suchembodiments, a user can store his/her settings, preferences, and/orchromatography data and results on a flash memory card to allow the userto essentially be “identified” by the controller 104, and for thechromatography system to be customized to each user.

The controller 104 can include chromatography software having agraphical user interface 106 to allow a user to interact with thecontroller 104 to manipulate and control the chromatography system 100.The graphical user interface allows a user to control a variety ofaspects of a chromatography run, including without limitation, changingparameters of a chromatography run, determining a mode of reportingchromatography results, determining a layout or format of reporting achromatogram or other chromatography results, and analyzingchromatography results. The details of the chromatography software andthe graphical user interface 106 will be described in greater detailbelow.

The controller 104 can be connected to the user interface 108. The userinterface 108 can include a variety of devices, including, withoutlimitation, one or more of the following: a keyboard 130, a mouse (notshown), a joystick (not shown), a touchpad 132 with a liquid crystaldisplay (LCD) screen 134, and a monitor 136 for displaying the graphicaluser interface 106 and having a touch screen 138 (operated by a stylus140 or a user's fingertip), and the like. In some embodiments, as shownin FIG. 1, the keyboard 130 can be positioned on a keyboard tray 142that can be coupled to and movable with respect to the housing 102. Thekeyboard tray 142 can be positioned within a recess of the housing 102to allow the keyboard 142 to be stowed away when not in use, therebycontributing to the compact configuration of the integratedchromatography system 100.

The cartridge holder 118 is coupled to a side of the housing 102 toposition the cartridge 120 in an easily accessible position, and toposition the cartridge 120 in a substantially vertical orientation. Insome embodiments, the cartridge holder 118 is formed by a portion of thehousing 102, or is integrally formed with one or more portions of thehousing 102. In some embodiments, such as the embodiment illustrated inFIGS. 1-6, the cartridge holder 118 includes a cartridge holder housing117 positioned to conceal at least a portion of tubing used to fluidlycouple one or more of the pump assembly 110, the sample injector 116,the cartridge 120, and the detector 122.

A variety of cartridge holders 118 formed of varying materials andhaving a variety of shapes can be used to hold the cartridge 120. Forexample, in some embodiments, the cartridge holder 118 can include aUNIMOUNT™ cartridge bracket, available from Analogix, Inc., Burlington,Wis.

A variety of cartridges 120 can be used with the chromatography system100. Accordingly, the cartridge holder 118 is adjustable or replaceableto accommodate a wide array of cartridge styles, materials and sizes. Insome embodiments, reusable, non-disposable cartridges are used. Forexample, in some embodiments, the cartridge 120 is formed of glass ormetal. In some embodiments, such as the embodiment illustrated in FIGS.1-6, the cartridge 120 is disposable and formed of a plastic. In someembodiments, the cartridge includes a machine readable identificationtag 133, including, but not limited to, a barcode or a radio-frequencyidentification (RFID) tag. The machine readable identification tag 133can include information relating to one or more of the following: thesize (e.g., length, inner diameter, outer diameter, etc.) of thecartridge 120, mass of the stationary phase 121 inside the cartridge120, and the type of stationary phase 121 used (e.g., silica,silica-based stationary phases, alumina-based stationary phases, etc.,each of which can include irregularly-shaped particles orspherically-shaped particles).

The machine readable identification tag 133 can be added to the exteriorof the cartridge 120 (e.g., an adhesive sticker, code written in ink),or the machine readable identification tag 133 can be integrally formedwith the cartridge 120 (e.g., written on the outer surface of thecartridge 120 with laser writing). In some embodiments, thechromatography system 100 can be equipped with a reader (e.g., anoptical scanner or a RF receiver) to read the machine readableidentification tag 133 on the cartridge 120. In some embodiments,reading the machine readable identification tag 133 of a cartridge 120of interest automatically inputs the necessary data regardingcharacteristics of the cartridge 120 into the chromatography software,allowing the user to skip having to enter cartridge information duringprogramming or select the cartridge 120 to be used from a list. As aresult, the machine readable identification tag 133 can reduce usererror and speed up the chromatography programming operation.

As shown in FIGS. 1 and 4, the machine readable identification tag 133is positioned on a lower, front portion of the cartridge 120, but itshould be understood that the machine readable identification tag 133can be positioned anywhere on the cartridge 120. In some embodiments,the machine readable identification tag 133 is positioned on thecartridge 120 at a location that will be adjacent or in close proximityto the reader on the chromatography system 100 when the cartridge 120 ispositioned for the chromatography process, to allow the machine readableidentification tag 133 to be read substantially simultaneously withpositioning the cartridge 120 for a chromatography run.

With reference to FIG. 7, the pump assembly 110 is positioned in aforward section of the base portion 127 of the housing 102. The pumpassembly 110 includes the pump 114, which moves fluid (e.g., the mobilephase and the sample 101) in the fluid path 112 of the chromatographysystem 100. The pump 114 is controlled by the controller 104 to pump thefluid in the chromatography system 100 at a given flow rate. Inembodiments employing more than one solvent 115, the pump assembly 110also includes a mixing valve 113 for mixing the solvents 115. The one ormore solvents 115 can include, without limitation, at least one ofmethanol, ethanol, 2-propanol, acetonitrile, ethyl acetate,tetrahydrofuran, acetone, dichloromethane, chloroform, diethyl ether,toluene, hexane, heptane, iso-octane, and combinations thereof. Thesolvents 115 can be stored and fluidly coupled to the pump assembly 110in a variety of ways. By way of example only, the embodiment illustratedin FIGS. 1-6 includes containers 125 for storing the solvents 115. Thecontainers 125 are positioned on a tray, or platform, 131 coupled to anupper portion of the monitor 136. Standard tubing and fittings known tothose of ordinary skill in the art can be used to fluidly couple thecontainers 125 to the pump assembly 110.

As shown in FIG. 7, a first solvent 115 a enters the base portion 127 ofthe housing 102 via a first aperture 144 a and travels to a first inlet146 a of the mixing valve 113 via tubing 148 a. A second solvent 115 benters the base portion 127 of the housing 102 via a second aperture 144b and travels to a second inlet 146 b of the mixing valve 113 via tubing148 b. The mixing valve 113 of the embodiment illustrated in FIG. 7 is adual gradient mixing valve and includes a first valve 150 a and a secondvalve 150 b that control the amount of the first solvent 115 a and thesecond solvent 115 b, respectively, in the mobile phase. The valves 150a, 150 b shown in the embodiment illustrated in FIG. 7 includemagnetically-controlled solenoid valves, but it should be understoodthat a variety of valves can be used to control the amount of thesolvents 115 present in the mobile phase, including, without limitation,a pinch valve, a rotary selector valve, and the like.

The opening of each valve 150 a, 150 b can be electronically controlled,and therefore can occur substantially instantaneously. The closing ofeach valve 150 a , 150 b, however, can be at least partially dependenton the resistance provided by the solvent 115 a, 115 b, respectively,based on the flow rate, temperature and viscosity of the solvent 115 a,115 b. By employing two valves 150 a, 150 b, when the first valve 150 a,for example, is in an open position, the first valve 150 a allows thefirst solvent 115 a to enter the mixing valve 113 to be added to themobile phase of the chromatography system 100. When the mixing valve 113receives a signal from the controller 104 to add the second solvent 115b to the mobile phase, the second valve 150 b will be signaled to moveto an open position, and the second solvent 115 b will begin being addedto the mobile phase. At the same time, the first valve 150 a can besignaled to move to a closed position. The closing of the first valve150 a may be at least partially dependent upon the flow rate,temperature and viscosity of the first solvent 115 a, but any delay inclosing of the first valve 150 a will be substantially overridden by thesubstantially instantaneous opening of the second valve 150 b, and anydelay that may occur in closing the first valve 150 a should notsignificantly affect the desired concentration of the mobile phase. Theillustrated embodiment includes two solvents 115 a, 115 b, but it shouldbe understood that the mixing valve 113 would function similarly forembodiments employing more than two solvents 115.

After the first solvent 115 a and the second solvent 115 b have beenmixed in the mixing valve 113 to form the mobile phase of thechromatography system 100, the mobile phase exits the mixing valve 113via an outlet 152 and travels to a connector 154 via tubing 156. Theconnector 154 shown in FIG. 7 is a Y connector 154 and allows the mobilephase to be directed to a first pump head 158 a and a second pump head158 b via a first inlet tube 160 a and a second inlet tube 160 b,respectively. Each pump head 158 a, 158 b includes a positivedisplacement pump. The first pump head 158 a and the second pump head158 b are adapted to operate 180 degrees out of phase, such that thefirst pump head 158 a and the second pump head 158 b alternate to createa steady flow profile with reduced pulsation and accurate gradientformation in the mobile phase of the chromatography system 100. Thefirst pump head 158 a and the second pump head 158 b can be easily andquickly replaced, and function together in the pump 114 to produceprecise dynamic gradient formation in the mobile phase. In theembodiment illustrated in FIG. 7, the pump 114 includes a binarygradient positive displacement pump, and specifically, an INTELLIFLOW™low pulsation pump, available from Analogix, Inc., Burlington, Wis. Avariety of other pumps or pump heads can be operated in the mannerdescribed above to produce a steady flow profile in the mobile phasewith reduced pulsation, including, without limitation, a piston pump(e.g., a piston pump available from Fluid Metering, Inc., Syosset, NY,or a piston pump available from Diener Precision Pumps), a multiplepiston pump (e.g., a SERIES DELTA® multiple piston pump, available fromMicropump, Vancouver, Wash.), etc.

The mobile phase flows out an outlet 162 of the pump 114 to a pressuretransducer 164 via outlet tubing 166, before exiting the base portion127 of the housing 102 via a third aperture 168 in the base portion 127(see FIGS. 1, 2 and 5). The pressure transducer 164 monitors thepressure in the mobile phase to determine if the fluid pressure producedby the pump 114 is above or below a desired fluid pressure. The pump 114can then be paused or otherwise adjusted based on feedback received fromthe pressure transducer 164. For example, the pressure transducer 164can send information to the controller 104 regarding the fluid pressure.The controller 104 can include a program to automatically pause orotherwise adjust the pump 114 based on the fluid pressure, including,without limitation, when the fluid pressure exceeds a preset thresholdor a fixed safety threshold. When the controller 104 pauses the pump 114in response to the fluid pressure sensed by the pressure transducer 164,the pump 114 can remain paused until a user inactivates the pause. Forexample, a ‘Resume’ button can appear in the graphical user interface106, either in an existing window or in a new window, or a ‘Resume’button on the user interface 108 can be pressed.

As shown in FIG. 2, the cartridge holder housing 117 includes acartridge holder inlet 180 positioned on the back of the cartridgeholder housing 117 that is fluidly coupled (e.g., via tubing 181) to thethird aperture 168 in the base portion 127 of the housing 102. Thecartridge holder inlet 180, in turn, is fluidly coupled to the inlet 119of the cartridge 120, for example, by tubing positioned within thecartridge holder housing 117. The cartridge holder housing 117 furtherincludes a cartridge holder outlet 182, which is fluidly coupled to theoutlet 123 of the cartridge 120, for example, by tubing positionedwithin the cartridge holder housing 117. The cartridge holder outlet 182is, in turn, fluidly coupled to the detector 122, as described below.The cartridge holder 118 is shown by way of example only, but it shouldbe understood that the cartridge holder 118 can include a variety ofother cartridge mounting means to couple the cartridge 120 to thechromatography system 100. For example, in some embodiments, thecartridge holder 118 does not include a cartridge holder housing 117,and the fluid path 112 is not concealed from the user by any suchhousing.

As shown in FIGS. 2, 3, 5 and 6, the detector 122 includes a flow cell170 coupled to a rear portion 172 of the housing 102 with a C-clamp 173.While other means for coupling the flow cell 170 to the chromatographysystem 100 are possible, the location and coupling means of the flowcell 170 of the illustrated embodiment allow for facile maintenance andreplacement. However, other fasteners known to those of ordinary skillin the art can be used in lieu of the C-clamp 173 without departing fromthe spirit and scope of the present invention.

The rear portion 172 of the housing 102 houses power supply componentsand components of the controller 104. The detector 122 is positioneddownstream of the chromatography cartridge 120 in the fluid path 112.Accordingly, fractions of the sample 101 are moved by the pump 114 fromthe outlet 123 of the cartridge 120 to an inlet 174 of the flow cell170. As shown in FIG. 2, in the illustrated embodiment, the fractions ofthe sample 101 are moved by the pump 114 from the outlet 123 to thecartridge holder outlet 182 to the inlet 174 of the flow cell 170.

At the detector 122, the fluid path 112 is exposed to electromagneticradiation, and the amount of radiation absorbed by each of the fractionspassing through the detector 122 is recorded and used to distinguish thefractions from one another. The fractions of the sample 101 exit thedetector 122 via an outlet 176 of the flow cell 170, and continuedownstream to the fraction collector 124. The detector 122 includes atransmitter that transmits the radiation to the fluid path 112, and areceiver that receives the radiation that was not absorbed by thefraction of the sample 101 passing through the detector 122 (i.e.,transmitted through the sample 101). The receiver sends data to thecontroller 104 regarding the amount of radiation transmitted througheach fraction. The amount of radiation absorbed by each fraction of thesample 101 is calculated, and the absorbance of each fraction is thendisplayed, usually in graphical form, on the graphical user interface106. The displayed absorbance data can then be manipulated and analyzedusing the chromatography software and graphical user interface 106, asdescribed below.

A variety of detectors 122 can be used to identify and distinguish thefractions, including, without limitation, a UV detector (e.g., anINTELLIUV™ detector, available from Analogix, Inc., Burlington, Wis.),an X-ray detector, an infrared detector, a visible light detector, arefractive index detector (e.g., a refractive index detector availablefrom LabAlliance), a photodiode array (PDA) UV detector, an evaporativelight scattering detector (ELSD) (e.g., an ELSD available from AlltechAssociates), and the like. In the embodiment illustrated in FIGS. 1-6,the detector 122 includes a fixed UV detector that emits UV radiationhaving a fixed wavelength. For example, in some embodiments, thedetector 122 emits UV radiation having a fixed wavelength of about 254nm. In some embodiments, the detector 122 includes a variable UVdetector that emits a UV radiation having a range of wavelengths. Forexample, in some embodiments, the detector 122 emits UV radiation havinga wavelength ranging from about 200 nm to about 300 nm.

After the fractions of the sample 101 have been detected by the detector122, peak detecting software identifies and distinguishes the differentpeaks, and determines a boundary between each “peak” or “non-peak”material in the sample 101. As used herein, the term “fraction” is usedto refer to “peak” material, or portions of the sample 101 that a usermay wish to detect and/or collect. In some embodiments, the peakdetecting software is a peak detecting module within the controller 104.In some embodiments, the peak detecting software communicates withfraction collecting software. The fraction collecting software controlsthe fraction collector 124, which expels the fractions into one or morecollection vessels 126, depending on the algorithm used to distinguishpeaks in the peak detecting software, and on user-prescribed settings.

The peak detecting software tells the fraction collecting softwarewhether fluid in the fluid path 112 is “peak” material or “non-peak”material, and whether it should be collected or not. In someembodiments, the peak detecting software functions independently andsimply identifies and distinguishes “peak” material from “non-peak”material, and maybe outputs this information to another instrument orother software, but does not control fraction collection. However, inthe illustrated embodiment, the peak detecting software communicateswith fraction collecting software, and thus, the terms “peak detectingsoftware” and “fraction collecting software” will be usedinterchangeably herein.

In some embodiments, dual, or multiple, wavelengths can be used in thedetector 122 to detect fractions in the sample 101. In some embodiments,dual, or multiple, types of detection (e.g., one UV and one ELSD, etc.)can be used to detect fractions. For example, if two wavelengths (e.g.,“colors”) of UV detection light are used, two chromatograms will becreated and displayed, showing how the fractions of the sample 101respond to the radiation source in the detector 122. The peak detectingsoftware can then use both chromatograms to distinguish “peak” materialfrom “non-peak” material. A variety of algorithms can be used toaccomplish this. For example, in some embodiments, the data points inone chromatogram may be weighted more heavily than data points inanother chromatogram. In some embodiments, the peak detecting softwaremay detect a new peak whenever a peak-triggering event occurs in any ofthe chromatograms. The peak detecting software can then output thisinformation to the fraction collecting software to signal to thefraction collector 124 when to collect a fraction and when to send it towaste. If more than one type of detector 122 is used, the delay volumebetween detectors 122 would need to be taken into account to synchronizethe chromatograms, so that data points relating to the same fraction inthe sample 101 can be correlated. The delay volume between subsequentdetectors can be entered into or calculated given the size (e.g., lengthand cross-sectional diameter or area) of tubing connecting an outlet ofa first detector 122 and an inlet of a second detector 122, and so on.

As best shown in FIG. 1, the fraction collector 124 includes an arm 192movable along a track 194 in an x-direction, the arm 192 having a lengthin a y-direction, substantially perpendicular to the x-direction. Thefraction collector 124 further includes a carriage 196 coupled to thearm 192 and movable along the length of the arm 192 in the y-direction.The carriage 196 includes a downwardly-directed nozzle 198 from whichthe one or more fractions of the sample 101 can be expelled into one ormore collection vessels 126. The arm 192 and carriage 196 allow thenozzle 198 to be positioned in an x-y plane above the collection vessels126. The carriage 196 can further include z-positioning means for movingthe nozzle 198 in a z-direction, orthogonal to the x-y plane, toward andaway from the collection vessels 126. Other types of carriages or gantrysystems having movement in one or more of the x-direction, they-direction and the z-direction can be used to position the nozzle 198over a desired collection vessel 126 for expulsion of one or morefractions of the sample 101, without departing from the spirit and scopeof the present invention.

As shown in FIGS. 1 and 4, the fraction collector 124 includes a divertvalve 184 that includes an inlet 186 that is fluidly coupled to theoutlet 176 of the flow cell 170 of the detector 122. Depending on thecollection mode of the fraction collecting software, which will bedescribed below, the fraction collecting software determines whether thefraction of the sample 101 is sent to a first outlet 188 of the divertvalve 184, which is fluidly coupled to waste, or a second outlet 190 ofthe divert valve 184, which is fluidly coupled to the nozzle 198. As aresult, fractions that are not to be collected will be directed to thefirst outlet 188 of the divert valve 184 to waste, and fractions thatare to be collected will be directed to the second outlet 190 of thedivert valve 184 and dispensed out of the nozzle 198 of the fractioncollector 124 and into a collection vessel 126.

In some embodiments, a user may wish to perform thin layerchromatography (TLC) with the contents of a collection vessel 126 byextracting a sample from a collection vessel 126 of interest andspotting the sample onto a TLC plate. In some embodiments, this isperformed manually. In some embodiments, the fraction collector 124 canbe programmed to automatically spot a TLC plate with each new fractionfrom the sample 101 that is about to be added to a new collection vessel126 (or at the very end of a collection). Alternatively, the user caninstruct the fraction collector 124, when desired, to spot a TLC platewith a desired fraction. In such embodiments, the carriage 196 of thefraction collector 124 can include mobility in the X-Y plane independentof the arm 192 to move away from the arm 192 to a TLC plate. Inaddition, the carriage 196 can include mobility in the z-direction tomove toward and away from a TLC plate.

In some embodiments, the user can select to perform TLC with at least aportion of the contents of each collection vessel 126 after achromatography run has completed. In some embodiments, the user canselect to perform TLC with at least a portion of one collection vessel126 corresponding to each detected fraction (i.e., some fractions canrequire more than one collection vessel 126), such as the firstcollection vessel 126 of a particular fraction, a middle collectionvessel 126 of a particular fraction, the last collection vessel 126 of aparticular fraction, the first and last collection vessels 126 of aparticular fraction, etc. In some embodiments, the user can select toperform TLC with at least a portion of the collection vessel 126corresponding to a fraction having the highest detected concentration(i.e., highest absorbance units measured by the detector 122). In someembodiments, the user can select to perform TLC with at least a portionof the collection vessel 126 corresponding to a fraction having thehighest or lowest slope of absorbance detected by the detector 122(i.e., the greatest increase or decrease in detected concentration).

As mentioned above, the fractions of the sample 101 can be collected inone or more collection vessels 126 positioned in one or more collectionvessel stands 128. In some embodiments, as shown in FIGS. 1, 2 and 5,the collection vessel stand(s) 128 includes a machine readableidentification tag 199, including, but not limited to, a barcode, aradio-frequency identification (RFID) tag, a magnetically activatedidentification tag (e.g., as available from Omron Corporation), anelectrically conductive identification tag (e.g., an IBUTTON® B serieselectrically conductive identification tag, available from DallasSemiconductor), etc. The machine readable identification tag 199 caninclude a variety of information, including, but not limited to: thesize of the collection vessel stand 128; the size of collection vessels126 stored in the collection vessel stand 128; the number of collectionvessels 126 the collection vessel stand 128 can hold; data relating tothe contents of the collection vessels 126 in the collection vesselstand 128; data identification such that a chromatogram relating to thecontents of the collection vessel stand 128 can be accessed by readingthe machine readable identification tag 199; and a globally uniqueidentifier (GUID) to definitively identify the collection vessel stand128 from all others. A GUID allows the chromatography system 100 and anyother instrument that may use or analyze the fractions of the sample 101to be able refer to a common database, so that one instrument can addinformation to the collection vessel stand 128 identified via the GUID,and another can access the information, and vice versa.

The machine readable identification tag 199 can be added to thecollection vessel stand 128 (e.g., an adhesive sticker, code written inink, a physical tag coupled to the collection vessel stand 128 by avariety of fasteners, including, without limitation, one or more of ascrew, nail, bolt, snap-fitting, press-fitting, and the like), or themachine readable identification tag 199 can be integrally formed withthe collection vessel stand 128 (e.g., written on the outer surface ofthe cartridge 120 with laser writing). In some embodiments, thechromatography system 100 can be equipped with a reader (e.g., anoptical scanner, a RF receiver, a magnetic reader, an electricalconduction reader, etc.) to read the machine readable identification tag199. In some embodiments, the machine readable identification tag 199can be read manually by a user, and the appropriate number or otheridentifier can be entered (e.g., by typing, speaking, or selecting froma list).

The position of the machine readable identification tag 199 is shown inFIGS. 1, 2 and 5 by way of example only, but it should be understoodthat the machine readable identification tag 199 can be positionedanywhere on the collection vessel stand 128. In some embodiments, themachine readable identification tag 199 is positioned on the collectionvessel stand 128 at a location that will be adjacent or in closeproximity to the reader on the chromatography system 100 when thecollection vessel stand 128 is positioned in the recess 202 of thehousing 102, to allow the machine readable identification tag 199 to beread substantially simultaneously with positioning the collection vesselstand 128 for a chromatography run.

In some embodiments, reading or entering the machine readableidentification tag 199 of a collection vessel stand 128 of interestautomatically inputs the necessary data regarding characteristics of thecollection vessel stand 128 into the chromatography software, allowingthe user to skip having to enter collection vessel stand 128 informationduring programming or select the collection vessel stand 128 to be usedfrom a list. As a result, the machine readable identification tag 199can reduce user error and speed up the chromatography programmingoperation. Furthermore, because the chromatography programming softwarewill have information about the collection vessel stand 128 being used,the controller 104 can pause the pump 114 when the collection vesselstand 128 no longer includes an empty collection vessel 126 availablefor the fraction collector 124. When the controller 104 pauses the pump114, the pump 114 can remain paused until a user inactivates the pause.For example, a ‘Resume’ button can appear in the graphical userinterface 106, either in an existing window or in a new window, or a‘Resume’ button on the user interface 108 can be pressed.

In some embodiments, the collection vessel stand 128 is reusable andnon-disposable, and is formed of a material such as metal. In someembodiments, the collection vessel stand 128 is disposable and formed ofa material such as plastic. In some embodiments, the collection vesselstand 128 is pre-filled with collection vessels 126 to eliminate thetime-consuming process of filling the collection vessel stand 128. Forexample, in some embodiments, the collection vessel stand 128 caninclude a plurality of integrally-formed collection vessels 126 (e.g., amolded tray, a thermoformed tray, etc.) to eliminate having to fill thecollection vessel stand 128 with collection vessels 126, or have thecollection vessel stand 128 pre-filled with collection vessels 126. Insome embodiments, a plurality of collection vessels 126 can beintegrally formed and have dimensions compatible with the collectionvessel stand 128 to allow a plurality of collection vessels 126 to beloaded into a collection vessel stand 128 at once. For example, a stripof collection vessels 126 having a length compatible with a length ofthe collection vessel stand 128 can be used to fill one row in thecollection vessel stand 128 at a time.

In some embodiments, as shown in FIGS. 1-6, the type of collectionvessel 126 used is a test tube, and accordingly, the type of collectionvessel stand 128 used is a test tube rack. In the embodiment illustratedin FIGS. 1-6, the housing 102 includes a recess 202 defined between anupper portion of the controller 104 and a lower portion of the monitor136. The recess 202 defines a collection vessel stand holder 135 and isdimensioned to receive one or more collection vessel stands 128, shownin FIGS. 1-6 as test tube racks.

In some embodiments, the chromatography system 100 includes one testtube rack 128, which can be positioned in the recess 202. After thefractions of the sample 101 have been expelled into one or more testtubes 126 in the test tube rack 128, the test tube rack 128 can beremoved and replaced with a new test tube rack 128 that includes emptytest tubes 126 ready to be filled by additional fractions from thesample 101, or fractions from a new sample. However, the chromatographyprocess may have to be paused to allow for removal and/or replacement ofthe test tube rack 128.

In the embodiment illustrated in FIGS. 1-6, the chromatography system100 includes two test tube racks 128 a, 128 b that are positioned on topof the controller 104 in the recess 202. The two test tube racks 128 a,128 b are identical and essentially have the appearance of one splittest tube rack 128. For simplicity, the split test tube rack 128 will bedescribed as two test tube racks 128 a, 128 b herein. Each test tuberack 128 a, 128 b includes a handle 204 a, 204 b to allow a user tograsp the test tube rack 128 a, 128 b during placement or removal of thetest tube rack 128 a, 128 b from the recess 202. Each of the two testtube racks 128 a, 128 b includes a flat edge 206 a, 206 b. Each flatedge 206 a, 206 b can be placed in abutting relationship with the otherflat edge 206 b, 206 a of the adjacent test tube rack 128 b, 128 a,respectively. Employing two test tube racks 128 a, 128 b allows a userto remove or replace one test tube rack 128 a, 128 b while the othertest tube rack 128 b, 128 a, respectively, is being filled by thefraction collector 124. The process of replacing the test tube rack 128a, 128 b that is currently not in use can be repeated indefinitelywithout interrupting the chromatography process to collect a largenumber of fractions. The LCD screen 134 on the touchpad 132 can displayinformation relating to whether the test tube racks 128 a, 128 b arefull or empty, and when they can be replaced. Alternatively, thisinformation can be displayed on the monitor 136.

FIGS. 8-20 illustrate a variety of screen shots of the graphical userinterface 106 of the chromatography software of the chromatographysystem 100, which will now be described in detail. The chromatographysoftware can include a variety of software, including, for example, theINTELLIPEAK™ chromatography programming software described above. Asshown in FIGS. 8-20, the graphical user interface 106 of thechromatography software includes a wizard program 220 that guides a userthrough the process of creating a chromatography run for the sample 101of interest. The wizard program 220 includes a main window 221 thatincludes four screens: a ‘General’ screen 222 (see FIG. 8), a‘Detection’ screen 224 (see FIG. 14), a ‘Pump’ screen 226 (see FIG. 16),and a ‘Chromatogram’ screen 228 (see FIG. 17). The four screens 222,224, 226, 228 are separated in a tabular format in the main window 221,and each screen 222, 224, 226, 228 can be accessed by selecting thecorresponding tab at the top of the main window 221. It should be notedthat the wizard program 220 can include as few as one screen and as manyscreens as desired to accomplish a variety of tasks, such as thosedescribed in greater detail below.

The main window 221 includes a static region 227 to allow a portion ofthe window 221 to be visible on each screen 222, 224, 226, 228. Thestatic region 227 can include one or more of the following: a ‘SampleID’ field 229 to record (e.g., using the keyboard 130) a name or numberto identify the sample 101; a method file title field 231 to display themethod being used and/or created; a ‘Run’ button 230, which can beselected to begin a chromatography run in the chromatography system 100;and a lamp button 233 (e.g., identified by a light bulb icon, as shownin the illustrated embodiment), which illustrates when the radiationsource (e.g., a UV lamp) is powered on in the detector 122.

The ‘Sample ID’ field 229 includes a sample identity wizard button 208.Selecting the sample identity wizard button 208 opens a ‘Sample IDSettings’ window 210, as shown in FIG. 8B. The ‘Sample ID Settings’window 210 includes an ‘Enable Auto Sequencing’ checkbox 212, a ‘Prefix’field 214, a ‘Starting’ field 216, an ‘OK’ button 218 which can beselected when the user is satisfied with the sample identity settings,and a ‘Cancel’ button 219 if the user does not wish to change or enterany data in the ‘Sample ID Settings’ window 210. When the ‘OK’ button218 is selected, the user will be returned to the ‘General’ screen 222.FIG. 8B illustrates one embodiment of auto sequencing sample identities.In the embodiment illustrated in FIG. 8B, the auto-sequencing featureincludes a fixed portion of a sample ID and a variable orauto-incrementing portion of the sample ID. The user can enter a fixedportion of a sample identity in the ‘Prefix’ field 214, and can use the‘Starting’ field 216 to specify a number from which to begin counting orincrementing to create a sequence of related runs.

After powering up, the chromatography software generally defaults to the‘General’ screen 222 of the wizard program 220. FIG. 8 illustrates the‘General’ screen 222 according to one embodiment of the invention. Asshown in FIG. 8, the ‘General’ screen 222 includes a ‘New’ button 232.Selecting the ‘New’ button 232 creates a new method file and clears alldata to reset the functions of the chromatography system 100. In someembodiments, after selecting the ‘New’ button 232, a dialog box willappear to ask the user, “Do you want to clear your current methodsettings?” and will include a ‘No’ button and a ‘Yes’ button. Selectingthe ‘Yes’ button will require the user to either enter a new method orload an existing method file.

The ‘General’ screen 222 further includes a ‘Load’ button 234, which canbe selected to find and select a saved method file (e.g., a *.mth file)from a Windows CE™-based browsing window. The ‘Load’ button 234 includesa load drop-down menu button 236. Selecting the load drop-down menubutton 236 activates a list of a variety of loading menu options,including, without limitation, at least one of ‘Load Method,’ which, ifselected, loads an existing method from memory within the controller104, a removable memory storage device (e.g., a Compact Flash card, afloppy disk, a compact disk (CD), etc.) or another computer that isconnected to the controller 104 via a hard-wired or wireless connection;‘Load Last Run,’ which, if selected, loads the last method run on thechromatography system 100; and ‘Load Results File,’ which, if selected,displays a list of automatically saved results files (e.g., a *.alxfile).

The ‘General’ screen 222 further includes a ‘Save’ button 238, which canbe selected to archive method data to memory (e.g., as a *.mth file)using a Windows CE™-based browsing window. The ‘Save’ button 238includes a save drop-down menu button 240. Selecting the save drop-downmenu button 240 activates a list of a variety of saving menu options,including, without limitation, at least one of ‘Save Method,’ which, ifselected, performs the same command as selecting the Save button 238;and ‘Save Results,’ which, if selected, opens a browser window to,allowing a user to save the current chromatography run as a result. Insome embodiments, the chromatography software automatically saveschromatography runs that last greater than three minutes when the run iscompleted.

The ‘General’ screen 222 further includes a ‘Description’ field 242,which includes a text field 244. A user can annotate the chromatographyrun by adding text to the text field 244. In some embodiments, the textin the text field 244 will automatically be saved with the method whenthe method is saved.

The ‘General’ screen 222 further includes a ‘Cartridge Type’ field 246,which allows a user to choose the type of chromatography cartridge 120to be used in the chromatography run. The chromatography cartridge 120can be chosen based on the size/amount of the sample 101 of interest anda retention factor (R_(F)) value. The ‘Cartridge Type’ field 246includes a cartridge drop-down menu button 248, which, when selected,provides a list of a variety of cartridges 120 available to the user. Byselecting a ‘System Options . . . ’ button 270, described below, theuser can navigate to the location of a cartridge configurable text fileto allow the user to build and modify the drop-down menu list forhis/her needs. The cartridges 120 can be listed by product name, productnumber, model name, model number, size of the cartridge 120 (e.g.,length, inner diameter, outer diameter, etc.), mass of the stationaryphase 121 in the cartridge 120, and combinations thereof.

The configurable text file can be in a table format, and can include avariety of fields, including, without limitation, at least one of IDnumber, cartridge volume (CV), equilibration time (e.g., in seconds),equilibration flow volume (e.g., in mL), purify flow volume (e.g., inmL), minimum flow volume (e.g., in mL), maximum flow volume (e.g., inmL), minimum sample size (e.g., in mg), maximum sample size (e.g., inmg), purge time (e.g., in seconds), maximum pressure able to withstand(e.g., in psi), inside diameter, outside diameter, length, suggestedflow rate (or minimum and maximum suggested flow rates), cross-sectionalarea, display description (i.e., how the cartridge will be identified inthe drop-down menu), comments, product family name, product name, etc.Other computations can be performed using data from one or more of thesefields. For example, in some embodiments, as shown in FIG. 8, after acartridge 120 is selected from the list (e.g., RS-40, as shown in FIG.8), a suggested flow rate and sample size for the cartridge 120 selectedwill appear in the ‘Cartridge Type’ field 246 (e.g., 25-50 mL/min. and30-1700 mg, as shown in FIG. 8), based on the data listed in thecorresponding fields for the cartridge selected.

The ‘General’ screen 222 further includes a ‘Solvents’ field 250, whichallows selection and identification of the solvents 115 to be used. The‘Solvents’ field 250 includes a first solvent field 252, named ‘A’ inFIG. 8, and a second solvent field 254, named ‘B’ in FIG. 8. Each of thefirst and second solvent fields 252, 254 includes a solvent drop-downmenu button 256, which, when selected, provides a list of availablesolvents that a user can select. In embodiments employing more than twosolvents 115, the Solvents field 250 includes a solvent field for everysolvent 115 to be used. In some embodiments, the user can choose asolvent from the list, or the user can enter a new solvent name in atleast one of the first solvent field 252 and the second solvent field254 using the keyboard 130. In some embodiments, a newly entered solventname can be stored when the method is saved. In some embodiments, anewly entered solvent name can be saved when a run is initiated forlonger than a predetermined amount of time (e.g., 3 minutes). The‘Solvents’ field 250 further includes a ‘Prime Pumps . . . ’ button 258.The ‘Prime Pumps . . . ’ button 258, when selected, opens a ‘PrimePumps’ window 260, as shown in FIG. 9.

With reference to FIG. 9, the ‘Prime Pumps’ window 260 includes a ‘FlowRate’ field 262 and a ‘Volume’ field 264 with incrementing buttons 266.A user can set a flow rate and volume for each solvent 115 to be used byusing the keyboard 130 to type in the ‘Flow Rate’ field 262 and the‘Volume’ field 264, or by using the incrementing buttons 266 to increaseor decrease the values displayed. A user can instead select a ‘UseDefault Values’ button 267 to select system default settings, which canbe stored in a configurable text file. The configurable text file can beaccessed by selecting the ‘System Options . . . ’ button 270.Alternatively, in some embodiments (not shown), one of the screens 222,224, 226, 228 (e.g., the ‘General’ screen 222) can include a button thatdirects the user to a ‘Cartridge Editor’ screen, which will allow theuser to edit the configurable text file using some type ofword-processing program (e.g., MICROSOFT® Windows NOTEPAD™ wordprocessing application). A user can also select one or more primingbuttons 268 to prime the first pump head 158 a (i.e., by selecting the‘Prime Pump A’ button 268 a), the second pump head 158 b (i.e., byselecting the ‘Prime Pump B’ button 268 b), or both (i.e., by selectingthe ‘Prime Pumps A & B’ button 268 c). When the flow rates and volumesfor the solvents 115 have been set, and the desired pump heads 158 a,158 b primed, the user can select a ‘Done’ button 269 to return to the‘General’ screen 222.

With continued reference to FIG. 8, the ‘General’ screen 222 furtherincludes the ‘System Options . . . ’ button 270. When the ‘SystemOptions . . . ’ button 270 is selected, a ‘System Options’ window 272 isopened, as shown in FIGS. 10-13B. The ‘System Options’ window 272 allowsa user to select a variety of settings and defaults in thechromatography system. With reference to FIGS. 10-13B, the ‘SystemsOptions’ window 272 includes five screens that are separated in atabular format: a ‘System’ screen 274, a ‘Solvents’ screen 276, a‘System Info’ screen 278, a ‘Control Panel’ screen 280, and a‘Calibration’ screen 281. Each screen 274, 276, 278, 280, 281 includes a‘Save & Close’ button 282. Selecting the ‘Save & Close’ button 282 willsave the selections chosen and return to the ‘General’ screen 222.

As shown in FIG. 10, the ‘System’ screen 274 includes a ‘Units’ field430, which allows the selection of pressure units in the system (e.g.,psi, as shown in FIG. 10), and includes a ‘Demo’ checkbox 432. Checkingthe ‘Demo’ checkbox 432 creates a chromatogram based on data pointsstored in a file, and does not actually run the pump assembly 110 or thedetector 122, but will operate the fraction collector 124, and allows auser to demonstrate how the chromatography system 100 works, withoutactually running a sample 101.

The ‘System’ screen 274 further includes an ‘Options’ field 434, where avariety of checkboxes 436 can be selected or deselected depending onuser preferences. For example, the user can select a ‘Use rack specifiedin the Method’ checkbox to store which collection vessel stand 128 theuser wishes to use with the method file (*.mth) created, so that eachtime that method file is loaded, the collection vessel stand 128settings are already in place. In addition, a ‘Show current timeindicator’ checkbox allows a user to select whether a vertical line willbe shown in a chromatogram that clearly shows the point in timecorresponding with each data point in the chromatogram.

The ‘System’ screen 274 further includes a ‘Delay Volume’ field 438,which allows for selection of a tubing size (e.g., cross-sectionaldiameter or area) and a length between the outlet 176 of the flow cell170 of the detector 122 and the inlet 186 of the divert valve 184 of thefraction collector 124. The cross-sectional area and length of tubingpositioned between the inlet 176 and the outlet 186 can allow forcalculation of a delay volume, which is explained in greater detailbelow.

The ‘System’ screen 274 can further include a ‘UV Lamp’ field 440, whichincludes an ‘Auto Shutoff (min)’ text field 442, which allows a user toenter or select an amount of time the detector lamp (UV or otherwise)will remain powered on before it automatically is powered off by thecontroller 104. The ‘UV Lamp’ field 440 further includes an ‘Auto-zeroruns’ checkbox 444, which can be selected to signify that the detectorshould be zeroed before each chromatography run.

As shown in FIG. 11, the ‘Solvents’ screen 276 includes a list ofsolvent names (and their relative strengths (e.g., polarity) shown inparentheses after the solvent name). Selecting the ‘Solvents’ screen 276in the ‘System Options’ window 272 provides access to this list ofsolvents, which may be edited in the ‘Solvents’ screen 276, or which maybe edited in a configurable text file that the ‘Solvents’ screen 276accesses. Editing the list of solvent names changes what is displayed inthe drop-down menus 256 in the ‘General’ screen 222. The list of solventnames and/or the configurable text file can be in a table format, and inaddition to a list of solvent names, can also include solventproperties, including, without limitation, at least one of density,boiling point, melting point, molecular weight, water solubility,chemical formula, vapor pressure, vapor density, etc.

As shown in FIG. 12, the ‘System Info’ screen 278 displays a variety ofinformation about the chromatography system 100, including the versionof the chromatography system 100 and/or software being used, copyrightinformation, which version of firmware for the pump 114 is being used,and the remaining hours of operation for the radiation source in thedetector 122. It should be understood that the ‘System Info’ screen 278can be used to display a variety of other parameters of thechromatography system 100.

As shown in FIG. 13, the ‘Control Panel’ screen 280 includes a varietyof Windows control panel icons, which, when selected (e.g., bydouble-clicking on the icon), allow access to a variety of Windowsfeatures. For example, selecting a ‘Windows Explorer’ icon 446 allowsthe user to access a Window desktop. By way of further example,selecting a ‘Stylus’ icon 448 opens a calibration window for calibratingthe touch screen 138. Other standard Windows control panel functionicons can be added to the ‘Control Panel’ screen 280 without departingfrom the spirit and scope of the present invention.

As shown in FIG. 13B, the ‘Calibration’ screen 281 includes a ‘RackOffset’ field 450 and a ‘Pump’ field 452. The ‘Rack Offset’ field 450allows a user to adjust the position of the fraction collector 124relative to collection vessels 126 positioned in a collection vesselstand 128 (i.e., perform x-y calibration of the fraction collector 124),by changing the left-right position of the fraction collector 124 andthe back-front position of the fraction collector 124 to prevent loss ofany portion of the sample 101, and to ensure that any fraction dispensedfrom the second outlet 190 of the divert valve 184 of the fractioncollector 124 is adequately collected in a collection vessel 126. Such acalibration may be necessary for a variety of reasons. For example, ifthe collection vessels 126 and/or the collection vessel stand 128 usedare slightly outside of their manufacturing tolerances, or includeslight defects, the x-y calibration of the fraction collector 124 may benecessary.

The ‘Pump’ field shows a volume per resolution value for the pump 114connected to the chromatography system 100. In some embodiments, theuser can enter or select this value. The user can then select a‘Calibrate’ button 454, which sends information to firmware in the pump114 to tell the pump 114 its volume per revolution. In some embodiments,firmware from the pump 114 tells the controller 104 what its volume perrevolution is, and this value is displayed.

FIG. 14 illustrates the ‘Detection’ screen 224 according to oneembodiment of the invention. The ‘Detection’ screen 224 includes a‘Collection Mode’ field 300, a ‘Threshold (AU)’ field 302, a ‘SlopeSelectivity’ field 304, a ‘Sample Savers’ field 306, and a ‘RackSettings’ field 308.

The ‘Collection Mode’ field 300 includes four fraction collection modeoptions, one of which can be selected by a user: a ‘Collect All’ mode310, a ‘Slope’ mode 312, a ‘Threshold’ mode 314, and a ‘Threshold &Slope’ mode 316. In some embodiments, a ‘Threshold or Slope’ mode 315,as shown in FIG. 14B, can be selected. The ‘Collect All’ mode 310 allowsall of the fractions of the sample 101 to be collected without beingseparated, so that the sample 101 is analyzed by the chromatographyprocess and expelled to the collection vessels 126 afterward.

The ‘Slope’ mode 312 uses the slope of a curve on a chromatogram 318(described below) of the fractions to determine when to separate thefractions. For example, if the slope of the curve decreases below acertain value, it may signal a break, or a boundary, between adjacentfractions, so the fraction collecting software will separate thefraction after the slope decreases below the predetermined amount. Thecontroller 104 and/or the fraction collecting software can then signalthe fraction collector 124 to stop dispensing the fraction to acollection vessel 126 and begin dispensing to waste. By way of furtherexample, if the slope of the curve increases again above a certainvalue, it may signal the beginning of a new fraction, and the controller104 and/or fraction collecting software can signal the fractioncollector 124 to stop pumping and switch the divert valve 184 to a“Drain” or “Waste” position (i.e., corresponding with the first outlet188), and move to a new collection vessel 126. The divert valve 184 canthen be switched back to a “Collect” position (i.e., corresponding withthe second outlet 190) when the carriage 196 is positioned over a newcollection vessel 126, and the pumping can be resumed.

In some embodiments, the ‘Slope’ mode 312 monitors the incoming signalfrom the detector 122 as discrete data points. The slope (in AU/min) iscalculated between adjacent points. The slope at each new incomingdiscrete data point is compared to a collection of previous data points(e.g., a rolling average, etc.) to determine if the slope at the newdata point passes a comparison test (e.g., if the slope at the newincoming data point exceeds that of the previous six points by a certainamount, etc.). In some embodiments, a plurality of tests can beperformed for each new data point, and the tests can be weighted tocalculate a total score for the new data point. The total score can becompared to some threshold value to determine if the new data point ispart of a continuing peak, if the new data point is going out of a peak,or if the new data point is part of a new peak, etc.

In some embodiments, the ‘Slope’ mode 312 computes the slope between afirst new data point and a prior data point, and stores this as ‘Slope1,’ for example. The ‘Slope’ mode 312 then computes the slope between asecond new data point and the first new data point, and stores this as‘Slope 2.’ Slope 2 and Slope 1 are then compared to determine if the newdata point is part of a continuing peak, going out of a peak, or part ofa new peak, etc.

The ‘Slope Selectivity’ field 304 can be used to adjust how the fractioncollecting software will select peaks or ignore peaks. The ‘SlopeSelectivity’ field 304 includes a sliding indicator 317 that can bemoved between ‘More’ and ‘Less’ selective positions on an axis 319.Moving the sliding indicator 317 along the axis 319 to different stepsalong the axis 319 chooses a different set of parameters for varioustests that are performed for peak selection based on slope. Accordingly,if a user wishes to collect more fractions (i.e., be “less selective”),he/she can slide the sliding indicator 317 using the stylus 140 or afingertip to a lower position on the axis 319 to collect more fractions,even those having subtle peaks on the chromatogram. Accordingly, thefraction collecting software will signal the fraction collector 124 tosend more fractions of the sample 101 to the collection vessels 126, andfewer to waste. On the other hand, if a user wishes to collect fewerfractions (i.e., be “more selective”), he/she can slide the slidingindicator 317 to a higher position on the axis 319 to collect only thefractions having well-defined peaks on the chromatogram. Accordingly,the fraction collecting software will signal the fraction collector 124to send fewer fractions of the sample 101 to the collection vessels 126,and more to waste. A peak display 321 illustrates an example of a peakcorresponding to each position along the axis 319.

In some embodiments, the ‘Slope Selectivity’ field 304 is replaced witha ‘Slope Sensitivity’ field 304 b, as shown in FIG. 14B, and the slidingindicator 317 b is movable along the axis 319 b, which is defined inunits of absorbance units (AU)/min. In some embodiments, as shown inFIG. 14B, the axis 319 b runs from a lower position of 0.005 AU/min. toan upper position of 1.000 AU/min., in increments of 0.005 AU/min.

The ‘Threshold’ mode 314 uses a baseline threshold value to determine ifa fraction is worth collecting or should be dispensed to waste. Forexample, if a portion of the curve of the chromatogram representing thesample 101 is below a threshold value (in absorbance units (AU)), thecontroller 104 and/or the fraction collecting software will direct thefraction collector 124 to dispense the fraction of the sample 101corresponding to that portion of the curve to waste. By way of furtherexample, if a portion of the curve of the chromatogram is above athreshold value of absorbance units, the controller 104 and/or thefraction collecting software will direct the fraction collector 124 todispense the fractions to the collection vessels 126 until the curvegoes below the threshold value again. Thus, any fraction of the sample101 corresponding to a portion of the chromatogram that is above apredetermined threshold of absorbance units will be collected. However,the fractions will be collected together, and will not be separated. Thethreshold value, in absorbance units (AU), can be specified in the‘Threshold (AU)’ field 302. A user can use the keyboard 130 to type avalue into a threshold text field 318, or a user can use incrementingbuttons 320 to increase or decrease the value displayed in the textfield 318. In some embodiments, the threshold value defaults to 0.100AU.

The ‘Threshold & Slope’ mode 316 includes a combination of the ‘Slope’mode 312 and the ‘Threshold’ mode 314. The ‘Threshold & Slope’ mode 316will only collect fractions that correspond to portions of thechromatogram that are above a predetermined threshold value, and thefraction collector 124 will separate the fractions and dispense eachfraction into a new collection vessel 126. The fraction collectingsoftware will separate the fractions based on the slope of thechromatogram, as described above.

The ‘Threshold or Slope’ mode 315 determines that fractions will becollected if the parameters for the ‘Threshold’ collection mode are metor if the parameters for the ‘Slope’ collection mode are met.

The ‘Sample Savers’ field 316 includes three checkboxes: a ‘CollectWaste in Tubes’ checkbox 322, a ‘Collect delay volume at front of peak’checkbox 324, and an ‘Auto-extend run during peak detection’ checkbox326. By checking the ‘Collect Waste in Tubes’ checkbox 322, the userspecifies that all “non-peak” portions of the sample 101 (i.e., portionsof the sample 101 that are not detected by the detector 122 as absorbingany radiation) will be collected, and not sent a waste container. Insome embodiments, collection vessels 126 can be identified as containing“waste” or “non-peak” material, and the fraction collector 124 can becontrolled to move to those collection vessels 126. For example, in someembodiments, collection vessels 126 for collecting “non-peak” materialcan be designated with black circles.

By checking the ‘Collect delay volume at front of peak’ checkbox 324,the user decides to collect a portion of a fraction corresponding towhen the detector 122 first noticed an increase in absorbance inradiation. In many cases, the most pure (i.e., most desirable) portionof a fraction of the sample 101 corresponds to a point in time where thedetector 122 first notices an increase in absorbance of radiation, whichcorresponds to a front of a peak on a chromatogram 370 (described belowin greater detail). Because a slope calculation needs more than one datapoint to determine the slope of the absorbance detected by the detector122, the slope can be calculated by comparing a new absorbance datapoint with a prior data point. If the ‘Collect delay volume at front ofpeak’ checkbox 324 is checked, and a sufficiently increasing slope(based on the settings of the fraction collecting software) has beenfound, the fraction collecting software determines where the slope wasfirst increasing by checking previous slope calculations, and controlsthe fraction collector 124 to collect a fraction beginning from thatpoint. Specifically, the “delay volume” refers to the volume of fluid inthe fluid path 112 between the outlet 176 of the flow cell 170 of thedetector 122 to the inlet 186 of the divert valve 184 on the fractioncollector 124. The delay volume can be calculated by knowing the lengthof tubing (or similar) used to connect the outlet 176 to the inlet 186,and the cross-sectional area of that tubing. In some embodiments, thechromatography system 100 is defaulted to send the delay volume at thefront of a peak that identifies a fraction to waste and continuecollecting a volume of fluid equal to the delay volume when collectingthe fraction at the back of the peak to ensure that the entire fractionis collected. By checking the ‘Collect delay volume at front of peak’checkbox 324, this default is overridden, and the delay volume is notsent to waste.

By checking the ‘Auto-extend run during peak detection’ checkbox 326,the user is telling the chromatography system 100 that if the fractioncollecting software has determined that the fraction collector 124should be collecting a fraction when the run time for the chromatographyrun expires, the run should continue, and the fraction collector 124should continue dispensing the fraction to a collection vessel 126 untilthe fraction collecting software determines that the entire fraction hasbeen collected, based on whichever collection mode is being used.

The ‘Rack Settings’ field 308 includes a ‘Current Rack:’ field 328,which displays the collection vessel stand 128 that is currentlypositioned in the recess 202 of the housing 102 for collecting fractionsof the sample 101. The ‘Rack Settings’ field 308 further includes a‘Change . . . ’ button 330. Selecting the ‘Change . . . ’ button opens a‘Rack Settings’ window 332, as shown in FIG. 15.

FIG. 15 illustrates one embodiment of the ‘Rack Settings’ window 332,which includes a spreadsheet 334 of collection vessel stand 128information and specifications and from which a user can highlight acollection vessel stand 128 of choice. A rack display 336 illustratesthe rack that is currently highlighted in the spreadsheet 334. The usercan add rows to the spreadsheet, edit the spreadsheet or delete rowsfrom the spreadsheet to suit his/her needs. Alternatively, in someembodiments, the collection vessel stand 128 selection can be completedin a manner similar to the manner in which the cartridge 120 and thesolvents 115 are selected, including a drop-down menu list that can bemodified by accessing a configurable text file. For example, in suchembodiments, a collection vessel stand configurable text file can beaccessed by selecting the ‘System Options . . . ’ button 270 on the‘General’ screen 222. Regardless of whether the collection vessel standoptions are listed in a spreadsheet or a configurable text file (whichcan be in a table format), the list can include a variety of datarelating to the collection vessel stands 128, including, withoutlimitation, at least one of the following: an ID no.; number of rows(i.e., if the collection vessel stand 128 is configured as a grid);number of columns; maximum number of collection vessels 126 that can beheld; position or distance data relating to the relative positions ofthe collection vessels 126 in the rack and/or the position of thecollection vessel stand 128 in relation to the recess 202 of the housing102 or the fraction collector 124; cap volume (e.g., in mL); displaydescription (i.e., the name given to the rack, which may be used in adrop-down menu, for example); and travel type (e.g., a “0” in a ‘traveltype’ field may refer to a serpentine travel path, meaning the fractioncollector 124 would increment through the collection vessel stand 128 bygoing down, over, up, over, down, and so on; a “1” may refer to a travelpath where the fraction collector 124 increments through the collectionvessel stand 128 by going down one column, down the next column, and soon; etc.).

The ‘Rack Settings’ window 332 further includes a ‘mL/Tube’ field 338,which allows the user to select a desired size of collection vessel 126,and a ‘Start Tube’ field 340, which allows the user to select whetherthe fraction collector 124 should begin dispensing fractions in thecollection vessel 126 positioned in the first position of the collectionvessel stand 128, or if the fraction collector 124 should begindispensing fractions in the first empty collection vessel 126.

The ‘Rack Settings’ window 332 further includes a ‘Near End of RackAlert’ field 342, which allows a user to program the chromatographysoftware to alert him/her when the fraction collector 124 is a certainnumber of collection vessels 126 from the end of what is available inthe collection vessel stand 128. The user can specify the number ofcollection vessels 126 prior to the end of the collection vessel stand128 he/she would like to be alerted. The alert can include anycombination of a variety of audible and visible alerts. Visible alertscan be displayed on one or more of the LCD screen 134 on the touchpad132, or the monitor 136.

The ‘Rack Settings’ window 332 further includes a ‘Close’ button 344,which can be selected when the user accepts or has completed theinformation in the ‘Rack Settings’ window 332. Selecting the ‘Close’button 344 will return to the ‘Detection’ screen 224.

FIG. 16 illustrates the ‘Pump’ screen 226 according to one embodiment ofthe present invention. The ‘Pump’ screen 226 includes an ‘Equilibration’field 346; a solvent gradient display chart 348; a solvent gradientspreadsheet 350 including an ‘Insert’ button 352 for inserting a rowinto the spreadsheet 350 and a ‘Delete’ button 354 for deleting a row inthe spreadsheet 350; a ‘Purification’ field 356, and a required solventvolume field 358.

The ‘Equilibration’ field 346 includes a ‘Time’ text field 360 and a‘Flow Rate’ text field 362, which allow a user to enter a time and flowrate, respectively, for equilibration of the chromatography system 100.An equilibration of the cartridge 120 will begin when a chromatographyrun is begun. Equilibrating the cartridge 120 and the chromatographysystem 100 is well-known in the art and therefore will not be discussedin greater detail below.

The solvent gradient spreadsheet 350 allows a user to define a solventgradient 364 for a chromatography run. By filling out a row in thesolvent gradient spreadsheet 350, the user enters a point in time in thechromatography run, an action at that point in time, and a percent ofone solvent 115 at that point in time. By entering several rows, a usercan define the relative amounts of the solvents 115 at given points intime throughout a chromatography run to define the solvent gradient 364,which is displayed in the solvent gradient display chart 348. Forexample, as shown in FIG. 16, solvent B is present at 10% for the firstthree minutes of the run, then increases linearly to 60% at thefifteen-minute mark of the run, and then increases linearly to 70% atthe seventeen-minute mark of the run.

The required solvent volume field 358 automatically updates based on thesolvent gradient 364 that is generated to tell the user how much of eachsolvent 115 will be required to complete the chromatography run. Therequired solvent volume field 358 also includes a cartridge volume (or‘column volume’; ‘CV’) flow rate field 366. As is well-known in the art,CV is the amount of mobile phase that fits in a packed cartridge 120.Accordingly, CV flow rate is a flow rate (e.g., CV/min.) based on theflow rate of the pump 114 (e.g., in mL/min), and the CV value associatedwith the selected cartridge 120 (e.g., data automatically entered afterselection of the cartridge 120 that can be stored in the cartridgesconfigurable text file). Some users may wish to work in CV rate, ratherthan pump flow rate. The CV flow rate field 366 tells how much time(e.g., in minutes) it will take for the mobile phase to pass through onecartridge 120, based on the CV flow rate.

FIG. 17 illustrates the ‘Chromatogram’ screen 228 according to oneembodiment of the present invention. The ‘Chromatogram’ screen 228displays a chromatogram 370 for a chromatography run as it is generatedduring the run, and after the run is completed. The ‘Chromatogram’screen 228 includes status bar 372 that displays the values for variousparameters at a given point in time. Some of the values displayed in thestatus bar 372 can be selected to be displayed in the status bar 372 ornot by selecting or deselecting corresponding checkboxes. The valuesthat can be displayed in the status bar 372 include absorbance units,time, flow rate, pressure, % second solvent 115 b, a numbercorresponding to which fraction of the sample 101 is being charted inthe chromatogram.

The status bar 372 further includes a ‘Threshold’ indicator 377, a‘Slope’ indicator 379, and a ‘Collecting’ indicator 381. Each indicator377, 379, 381 has three states: Green: triggered, Red: not triggered,and Gray: inactive. If the collection mode chose includes both of thethreshold mode and the slope mode, both corresponding indicators 377,379 will be active. The ‘Slope’ indicator 379 is triggered and turnsgreen when the chromatogram 370 begins to rise quickly enough, based onthe ‘Slope Selectivity’ parameters selected in the ‘Detection’ screen224. When the fraction collecting software has determined that the peakhas passed, the ‘Slope’ indicator 379 is no longer triggered and turnsto red. Similarly, the ‘Threshold’ indicator 377 turns green when thechromatogram 370 reaches or exceeds the threshold absorbance unit thatwas set in the ‘Threshold (AU)’ field 302 in the ‘Detection’ screen 224.Whenever the chromatogram falls below this threshold value, the‘Threshold’ indicator 377 turns red. The ‘Collecting’ indicator 381turns green when the fraction passing through the detector 122 will becollected, as determined by the fraction collecting software. The actualcollection is delayed until the fraction reaches the divert valve 184 ofthe fraction collector 124. The ‘Collecting’ indicator 381 will turngreen some time before the divert valve 184 switches to the secondoutlet 190.

The ‘Chromatogram’ screen 228 further includes a graphic rack display374 of the collection vessel stands 128 being used. The graphic rackdisplay 374 is dynamic and color-coded such that the fraction of thesample 101 in each collection vessel 126 in the collection vessel stand128 can be matched up with a similarly-colored color band 375 displayedin a lower portion of the chromatogram 370 to visually determine whichfraction is in which collection vessel 126. Such ‘fraction mapping’allows facile retrieval of a particular fraction of interest for storageor further analysis. Color-coding fraction mapping can be accomplishedusing a variety of software tools to match up the collection vessels 126with the corresponding portions of the chromatogram 370. For example,FRACTRAC™ fraction mapping software (available from Analogix, Inc.,Burlington, Wis.) can be used for this purpose. Other types of matchingor coding can be used in addition to color-coding without departing fromthe spirit and scope of the present invention, including, withoutlimitation, patterns, shading, etc.

As shown in FIG. 17, a fraction can take up more than one collectionvessel 126. In such embodiments, the user may wish to consolidate all ofthe collection vessels 126 pertaining to one fraction into onecontainer. In some embodiments, the chromatography system 100 allows forautomatic consolidation of fractions. This could be set to be performedautomatically after the completion of a run, or a user could instructthe chromatography system 100 (e.g., via the graphical user interface106) to consolidate the fractions. For example, after a run, the usercan instruct the chromatography system 100 to take all of the collectionvessels 126 pertaining to a first fraction and consolidate them into onelarger vessel. By way of further example, perhaps the user is notinterested in keeping a second fraction, so he/she can instruct that thecontents of all of the collection vessels 126 pertaining to the secondfraction be sent to waste, and so on. Finally, the user can dispose ofall of the used and empty collection vessels 126 (or, in someembodiments, the chromatography system may wash the collection vessels126 for reuse). Consolidation of the fractions by the chromatographysystem 100 can save the user from having to perform all of the liquidhandling steps manually, which can be tedious and time-consuming.

The ‘Chromatogram’ screen 228 further includes display manipulationbuttons 376. In some embodiments, the display manipulation buttons 376include toggle buttons 378 to control whether the status bar 372,chromatogram 370 and/or graphic rack display 376 are displayed. Thedisplay manipulation buttons 376 further include a chromatogram sizeadjustment button 380 that can increase or decrease the amount of the‘Chromatogram’ screen 228 that is taken up by the chromatogram 370. Thedisplay manipulation buttons 376 further include absorbance unit zoombuttons 382 for zooming in and out on the y-axis of the chromatogram370, and time zoom buttons 384 for zooming in and out on the x-axis ofthe chromatogram 370.

As described above, the static region 227 of the main window 221 isaccessible from any of the four screens 222, 224, 226, 228 and includesthe ‘Run’ button 230. As shown in FIG. 18, the first time the ‘Run’button 230 is selected, a ‘Run Summary’ window 290 will open to displaya summary of the method about to be run on the chromatography system100. In some embodiments, as shown in FIG. 14, the summary includes thetitle of the method file, the cartridge type, the collection vesselstand 128 chosen, the fraction collection mode chosen, and the solvents115 chosen. The ‘Run Summary’ window 290 can further include a‘Continue’ button 292 to begin the chromatography run (or to beginequilibrating the chromatography cartridge 120), an ‘Abort’ button 294to abort the chromatography run, and a ‘Do not show the Run Summaryagain’ checkbox 296. If the ‘Do not show the Run Summary again’ checkbox296 is checked, a chromatography run can be started by simply selectingthe ‘Run’ button 230, and the ‘Run Summary’ window 290 will not openprior to beginning the chromatography process.

If the ‘Continue’ button 292 is selected in the ‘Run Summary’ window290, an ‘Equilibrating Cartridge’ window 386 will open, as shown in FIG.19, and the chromatography system 100 will begin equilibrating thechromatography cartridge 120. The ‘Equilibrating Cartridge’ window 386includes a time progression bar 388 and a ‘Time Remaining’ text field390 to visually display the progress of equilibration. The‘Equilibrating Cartridge’ window 386 further includes a ‘Skip’ button392, which, when selected, allows the chromatography run to beginwithout completing equilibration, and a ‘Pause’ button 394, which, whenselected, allows at least one of the equilibration and thechromatography run to be paused. After the ‘Pause’ button 394 isselected, a ‘Resume’ button (such as the ‘Resume’ button 404 shown inFIG. 20, described below) and an ‘Abort’ button (such as the ‘Abort’button 406 shown in FIG. 20, described below) will appear, and the usercan then select whether he/she wishes to continue running thechromatography process, or whether he/she wishes to abort the run.

When the ‘Continue’ button 292 of the ‘Run Summary’ window 290, orsimply when the ‘Run’ button 230 is selected (e.g., when the ‘Do notshow the Run Summary again’ checkbox 296 has been checked in the ‘RunSummary’ window 290), the ‘Run’ button 230 changes to a ‘Pause’ button295, as shown in FIG. 19. The ‘Pause’ button 295 can be selected at anytime throughout the chromatography process to pause the run. By pausingthe run using the ‘Pause’ button 295, the user can edit any of themethod file and run settings “on-the-fly” before resuming thechromatography run. Such “on-the-fly” editing provides flexibility inthe chromatography process and eliminates errors and wastedchromatography runs by allowing the run to be paused and edited beforean error occurs. The ‘Pause’ button 295 in the graphical user interface106 is only one way to implement “on-the-fly” editing. Thechromatography run can be paused using a variety of other means,including, without limitation, a ‘Pause’ button on the touchpad 132,etc.

As shown in FIG. 20, an ‘Inject Sample’ window 396 will open when eitherthe ‘Skip’ button 392 is selected, or the cartridge 120 is finishedequilibrating. The ‘Inject Sample’ window 396 includes an ‘SI ModuleInjection’ instructions field 398 and a ‘Syringe Injection’ instructionsfield 400, which each include instructions for injecting the sample 101using the sample injector 116. Alternatively, as described above, thesample 101 can be loaded directly into the inlet 119 of the cartridge120.

The ‘Inject Sample’ window 396 further includes an ‘Auto-zero UVdetector at start of run’ checkbox 402. If the ‘Auto-zero UV detector atstart of run’ checkbox 402 is checked, the detector 122, whether it is aUV detector or another type of detector 122, will be zeroed (i.e., thebaseline absorbance reading will be zeroed) before proceeding with thechromatography run. The ‘Inject Sample’ window 396 further includes a‘Resume’ button 404 to continue with the chromatography run, and an‘Abort’ button 406 to abort the chromatography run.

FIG. 21 illustrates the touchpad 132 according to one embodiment of thepresent invention. The touchpad 132 includes the LCD screen 134, anumeric keypad 408, a power indicator 410, ‘Collect Now’ controls 412, a‘Zero Base’ button 414, a gradient hold button 416, a tube advancebutton 418, and a ‘Drain’ button 420.

The numeric keypad 408 can be used to enter numeric data into any of thetext fields in the wizard program 220, enter numeric data in or selectitems from menus in the LCD screen 134 on the touchpad 132, or enternumeric data in any other word processing or spreadsheet programassociated with the controller 104. The numeric keypad 408 includesbuttons enumerated from 0 to 9 and, a ‘CLEAR’ button for clearing out afield, and a ‘BKSP’ button for deleting the last character entered. Thepower indicator 410 indicates when the fraction collector 124 is poweredon.

The ‘Collect Now’ controls 412 include a Collect Now ‘Start’ button 422and a Collect Now ‘End’ button 424. The ‘Collect Now’ function allows auser to override peak detecting software and the absorbance datameasured by the detector 122 to collect a fraction that appearsdesirable or interesting based on the chromatogram, but which normallywould be sent to waste, based on the settings and the collection mode ofthe fraction collecting software. Pressing the Collect Now ‘Start’button 422 causes the controller 104 to send a signal to the fractioncollector 124, and particularly, to the divert valve 184 to send all ofthe fractions of the sample 101 (in some embodiments, after the delayvolume has passed through the divert valve 184) out the second outlet190 to the collection vessels 126 to be collected, no matter what datahas been acquired from the detector 122. Pressing the Collect Now ‘End’button 424 resumes the separation of the fractions of the sample 101based on the settings and the collection mode of the fraction collectingsoftware. The ‘Collect Now’ controls 412 can be used to make sure thatthe sample 101 does not go to waste, and can be used to override thefraction collecting software at the last minute, if it is determinedthat a mistake was made in programming the settings of the fractioncollecting software. In some embodiments, the ‘Collect Now’ controls 412include simply one toggle button that can be pressed once to activatethe ‘Collect Now’ function, and can be pressed a second time to disablethe ‘Collect Now’ function.

The ‘Zero Base’ button 414 allows a user to manually zero the absorbanceunit baseline in the detector 122. In some embodiments, fractions from aprevious sample may have been present in the flow cell 170 of thedetector 122 when the detector 122 was zeroed using the wizard program220 (i.e., by checking the ‘Auto-zero UV detector at start of run’checkbox 402, as shown in FIG. 20 and described above), causinginaccurate absorbance unit measurements. In other embodiments, the‘Auto-zero UV detector at start of run’ checkbox 402 may not have beenchecked, and the detector 122 may not have been zeroed between runs.Accordingly, the user can press the ‘Zero Base’ button 414 on thetouchpad 132 to zero the detector 122 after priming or flushing thefluid path 112. In still other embodiments, adding a solvent 115 that,even slightly, absorbs radiation in the detector 122 during thechromatography run may artificially increase the absorbance value. The‘Zero Base’ button can be pressed to compensate for this.

The gradient hold button 416 can be used to override the solventgradient 364 that was generated using the ‘Pump’ screen 226 of thewizard program 220. In some cases, the user may see a peak on thechromatogram 370 of a fraction that is being collected at a givensolvent concentration, and the user may wish to override the solventgradient 364 that was originally generated to continue collection ofthat peak. In that case, the user may press the gradient hold button 416to maintain a desired solvent concentration. When the user presses thegradient hold button 416 a second time, the solvent concentration willresume following the originally generated solvent gradient 364. Forexample, with respect to the solvent gradient 364 shown in FIG. 16, ifthe user determined at the 8-minute mark to hold the solventconcentration at about 30% of the second solvent 115 b (% B in FIG. 16),he/she would push the gradient hold button 416 at the 8-minute mark, andthe solvent gradient would follow a new curve, namely, a modifiedsolvent gradient 364 b. According to the modified solvent gradient 364 bshown in FIG. 16, the user pressed the gradient hold button 416 again atthe twelve-minute mark. After the gradient hold button 416 is pressed asecond time, the solvent concentration resumes following the generatedsolvent gradient 364 by increasing from 30% B to 50% B within 3 minutes,instead of 7 minutes (i.e., has a greater slope between the 12-minutemark and the 15-minute mark to “catch up” to the originally generatedsolvent gradient 364). As a result, the gradient hold button 416 canfunction as a manual override of the originally generated solventgradient 364.

The tube advance button 418 can be pressed to advance the fractioncollector 124, and particularly, the carriage 196 of the fractioncollector 124 to the next collection vessel 126. This can be used for avariety of purposes. In some embodiments, the tube advance button 418can be used as a safety measure if the user observes that a collectionvessel 126 may be about to overflow, for example, if the collectionvessel 126 was not completely empty when it began to be filled. In someembodiments, the tube advance button 418 can be pressed when the userobserves what appears to be a new peak in the chromatogram 370, butwhich is not being detected as a new peak by the fraction collectingsoftware, based on the collection mode and parameters that have beenset. In such embodiments, the user can press the tube advance button 418to begin collecting the new peak in a new collection vessel 126 to,essentially, manually separate the fractions of the sample 101.

The drain button 420 can be pressed to tell the fraction collector 124,and particularly, the divert valve 184 of the fraction collector 124 tosend all fractions of the sample 101 out the first outlet 188 to waste.When the drain button 420 is pressed, the fraction collecting softwareis overridden, and no fractions are collected until the drain button 420is pressed a second time. The drain button 420 can be pressed a secondtime to resume collecting fractions according to the settings in thefraction collecting software.

The LCD screen 134 on the touchpad 132 can be used to display currentsettings in the chromatography process relating to programming a run,priming the chromatography system 100, and/or running the chromatographysystem 100.

In operation, a user would power on the chromatography system 100 andwould open the chromatography programming software, and particularly,the wizard program 220. The wizard program 220 would default to the‘General’ screen 222, and the user would have the option of creating anew method file or loading an existing method file, as described above.The user can then select the type of cartridge 120 and solvents 115 tobe used for the chromatography run, and enter any notes into the textfield 244 of the ‘Description’ field 242. The user can change varioussystem options, as described above, by selecting the ‘System Options . .. ’ button 270, and the user can prime one or both of the pump heads 158a, 158 b and enter or select the flow rate and volume of the solvents115 by selecting the ‘Prime Pumps . . . ’ button 258. The user can entera sample identification into the ‘Sample ID’ field 229, or the user canenable auto-sequencing by selecting the sample ID wizard button 208, asdescribed above with respect to FIG. 8B.

The user can then advance to the ‘Detection’ screen 224 by clicking onthe ‘Detection’ tab at the top of the main window 221 of the wizardprogram 200. The ‘Detection’ screen 224 includes default settings, andmany users will not make adjustments to the ‘Detection’ screen 224,except to modify the ‘Rack Settings’ field 308. However, any of theother fields in the ‘Detection’ screen 224 can be modified, as describedabove.

The user can then advance to the ‘Pump’ screen 226 by clicking on the‘Pump’ tab at the top of the main window 221. The user can generate asolvent gradient 364 by adding rows to the solvent gradient spreadsheet350, based on the sample 101 of interest. The user can then modify the‘Equilibration’ parameters in the ‘Equilibration’ field 346 or the‘Purification’ flow rate in the ‘Purification’ field 356, as describedabove. The user can also make note of the required amount of solvents115, based on the display in the required solvent volume field 358.

The user can then advance to the ‘Chromatogram’ screen 228 by clickingon the ‘Chromatogram’ screen 228 to run and monitor the chromatographyrun. The ‘Chromatogram’ screen 228 can also be used to view previousresults. If the user wishes to begin a chromatography run, he/she canselect the ‘Run’ button 230 located in the static region 227 of the mainwindow 221. As mentioned above, unless the ‘Run Summary’ window optionhas been turned off, after the user has selected the ‘Run’ button 230,the ‘Run Summary’ window 290 will open and allow the user to review therun summary and abort beginning the run by selecting the ‘Abort’ button294, if desired, or continue the chromatography process by selecting the‘Continue’ button 292. If the ‘Continue’ button 292 is selected, thechromatography system 100 will begin equilibrating the cartridge 120, asshown in FIG. 19 and described above, which can be skipped or paused byselecting the ‘Skip’ button 392 or the ‘Pause’ button 394, respectively.In addition, the ‘Run’ button 230 has now changed to the ‘Pause’ button395, and the ‘Pause’ button 395 can be used to pause and restart the runor pause and abort the run.

After the cartridge 120 has been equilibrated, or after equilibrationhas been skipped, the ‘Inject Sample’ window 396 will open, and user canintroduce the sample 101 into the chromatography system 100 in two ways,as described above. After the sample 101 has been injected, the user canselect the ‘Resume’ button 404 to resume the chromatography run, or the‘Abort’ button 406 abort the chromatography run. If the user selects the‘Resume’ button 404, the pump assembly 110 will move the solvents 115from the containers 125 to the mixing valve 113 to be mixed according tothe solvent gradient 364 specified to form the mobile phase of thechromatography system 100, and pumped by the pump 114 through the fluidpath 112 and to the cartridge 120. The sample 101 will be moved throughthe cartridge 120 with the mobile phase of the chromatography system100, passed the stationary phase 121. The sample 101 will be separatedinto fractions based on the relative affinities of the fractions in thesample 101 for the mobile phase and the stationary phase 121. Thefractions will then be directed from the outlet 123 of the cartridge 120to the inlet 174 of the flow cell 170 of the detector 122, where thefractions of the sample 101 will be identified and distinguish.

Based on the reading from the detector 122, a chromatogram 370displaying the absorbance of radiation of each fraction will be createdin the ‘Chromatogram’ screen 228 of the wizard program 220 of thegraphical user interface 106. Based on the parameters and collectionmode chosen of the fraction collecting software, the fraction collectingsoftware will determine when one fraction ends and a new fractionbegins. As fractions pass through the flow cell 170, and out the outlet176 of the flow cell 170, the fractions are directed to the divert valve184 of the fraction collector 124, and either sent to waste via thefirst outlet 188, or sent out of the nozzle 198 to a collection vesselvia the second outlet 190, depending on the signals received from thecontroller 104, based on the fraction collecting software.

When a new fraction is detected, the arm and/or the carriage 196 of thefraction collector 124 will index to a new collection vessel 126 tocollect the newly identified fraction in a new collection vessel 126.The fraction collector 124 will also index to a new collection vessel126 when one collection vessel 126 is almost fall, based on the datastored in the chromatography programming software regarding the type ofcollection vessel 126 and collection vessel stand 128 being used. Asshown in FIG. 17, the graphic rack display 374 in the ‘Chromatogram’screen 288 will show the relationship (i.e., fraction mapping) betweenthe contents of the collection vessels 126 and the peaks of thechromatogram 370.

Various features and aspects of the invention are set forth in thefollowing claims.

1. A method for analyzing a sample of interest using chromatography, thesample including fractions, the method comprising: providing acontroller; providing a fluid path through which a mobile phasecomprising the sample moves during chromatography; moving the mobilephase through the fluid path in a chromatography run according tochromatography run parameters; sending a signal to the controller at anytime during the chromatography run to pause the chromatography run; andediting the chromatography run parameters.
 2. The method of claim 1,further comprising: separating the fractions of the sample, detectingthe fractions of the sample, and selectively collecting fractions of thesample.
 3. The method of claim 1, further comprising resuming thechromatography run after editing the chromatography run parameters. 4.The method of claim 1, wherein editing the chromatography run parametersincludes editing at least one of: the flow rate of the mobile phase, anymanually-entered information, a fraction collection mode, a fractioncollection threshold, a fraction collection slope sensitivity, a solventgradient of the mobile phase, and combinations thereof.
 5. The method ofclaim 1, wherein sending a signal to the controller to pause thechromatography run includes activating a feature in a graphical userinterface.
 6. The method of claim 1, wherein sending a signal to thecontroller to pause the chromatography run includes sending a signal toa pump to pause movement of the mobile phase in the fluid path.
 7. Themethod of claim 1, further comprising separating the fractions of thesample by moving a mobile phase comprising the sample through achromatography cartridge comprising a stationary phase.
 8. A method foranalyzing a sample of interest using chromatography, the methodcomprising: selecting a parameter for a chromatography run of thesample; commencing the chromatography run according to the selectedparameter; transporting the sample through a fluid path in achromatography system to perform the chromatography run; pausing thechromatography run; modifying the selected parameter of thechromatography run; and recommencing the chromatography run according tothe modified parameter.
 9. The method of claim 8, wherein the parameteris selected using a graphical user interface in communication with thechromatography system.
 10. The method of claim 8, further comprising,separating the sample into a plurality of fractions, detecting thefractions, and determining a composition of at least one fraction. 11.The method of claim 10, wherein separating the sample includes movingthe sample through a chromatography cartridge comprising a stationaryphase.
 12. The method of claim 10, further comprising at least partiallyfilling a vessel with a portion of the sample.
 13. The method of claim8, wherein modifying the selected parameter of the chromatography runincludes modifying at least one of a flow rate of the sample, anymanually-entered information, a fraction collection mode, a fractioncollection threshold, a fraction collection slope sensitivity, a solventgradient of the mobile phase, and combinations thereof.
 14. The methodof claim 8, wherein pausing the chromatography run includes instructinga pump in the chromatography system to pause movement of the sample inthe fluid path.
 15. A method for analyzing a sample of interest usingchromatography, the method comprising: accessing a chromatography systemto select a plurality of parameters of a chromatography run via agraphical user interface; commencing the chromatography run for thesample of interest; pausing the chromatography run via the graphicaluser interface; modifying at least one of the selected parameters of thechromatography run; recommencing the chromatography run according to theat least one modified parameter; separating the sample into a pluralityof fractions; and collecting at least one of the fractions in a vessel.16. The method of claim 15, wherein separating the sample into aplurality of fractions includes moving the sample through achromatography cartridge.
 17. The method of claim 15, wherein modifyingat least one of the selected parameters of the chromatography runincludes modifying at least one of a flow rate of the sample, anymanually-entered information, a fraction collection mode, a fractioncollection threshold, a fraction collection slope sensitivity, a solventgradient of the mobile phase, and combinations thereof.
 18. The methodof claim 15, wherein pausing the chromatography run includes instructinga pump in the chromatography system to pause movement of the samplethrough a fluid path in the chromatography system.